| Objective:The purpose of this study is to observe the changes of VEGF-Dll4/Notch signal pathway and the effect of electroacupuncture in the acute phase of cerebral infarction,and to enrich the molecular mechanism of acupuncture in the treatment of cerebral infarction from the perspective of promoting angiogenesis.Methods:Wistar rats were randomly divided into control group,model group and EA group according to the random number table method.The latter two groups were divided into 10 time groups according to 1h,3h,6h,9h,12 h,18h,24 h,3d,7d and 12 d after operation.The middle cerebral artery occlusion model was reproduced by the improved longa method.Neurological severity(NSS)scoring method was conducted in evaluating neural functional deficient degree of rats after MCAO.RT-PCR,Western blot and ELISA were used to detect the expression of VEGF and its receptor VEGFR-1,VEGFR-2,and the key factors of Dll4/Notch signaling pathway Notch 1,Notch 4,Jagged1,Dll4,hes1 mRNA and protein and the effect of EA intervention.Results:1 Interventional effect of EA on NSS scores: NSS scores of rats in control groups at each time point were 0.The NSS score of model group decreased gradually with the prolongation of ischemia time.The trends in EA groups were approximately equal to model groups,with lower degree in comparison with that of model groups at equivalent phases.Changes were statistically significant at 3d,7d and 12d(P<0.05,P<0.01)2 Interventional effect of EA on the expression of VEGF and its receptors VEGFR-1,VEGFR-2 mRNA and protein:(1)The mRNA and protein expression of VEGF:the mRNA expression at 3h～12d and protein expression at 6h～7d of rats in model groups was up-regulated significantly compared to that in control groups(P<0.05,P<0.01).The mRNA levels of rats in EA groups was higher than that of simultaneous model group in 3h～12d,and were superior to that in model groups at 3h,12h～24h(P<0.05,P<0.01).While the protein levels of rats in EA groups was higher than that of the model group at the same time at 3h～12d,and were superior to that in model groups at 3h,12 h and 18h(P<0.05).(2)The mRNA and protein expression of VEGFR-1:the mRNA expression at 3h～12d and protein expression at 9h ～ 3d of rats in model groups was up-regulated significantly compared to that in control groups(P<0.05,P<0.01).The mRNA levels of rats in EA groups was higher than that of the same phase model group in 3h～12d,and were superior to that in model groups at 3h,12 h and 18h(P<0.05,P<0.01).While the protein levels of rats in EA groups was higher than that of the model group at the same time at 3h～12d,and were superior to that in model groups at 12 h and 24h(P<0.05,P<0.01).(3)The mRNA expression of VEGFR-2:the mRNA expression at 3h～12d and protein expression at 6h～12d of rats in model groups was up-regulated significantly compared to that in control groups(P<0.05,P<0.01).The mRNA levels of rats in EA groups was higher than that in model group,and were superior to that in model groups at 3h,12 h and 24h～7d(P<0.05,P<0.01).While the protein levels of rats in EA groups was higher than that of the model group,and were superior to that in model groups at 9h,3d and 7d(P<0.05,P<0.01).3 Interventional effect of EA on the expression of Notch1,Notch4,Jagged1,Dll4,hes1 mRNA and protein related to Dll4/Notch signaling pathway:(1)The mRNA and protein expression of Notch1:the mRNA expression at 3h～12d and protein expression at 9h～12d of rats in model groups was up-regulated significantly compared to that in control groups(P<0.05,P<0.01).The mRNA levels of rats in EA groups was higher than that of the same phase model group in 12 h,24h～12d,and were superior to that in model groups at 3d and 7d(P<0.05,P<0.01).While the protein levels of rats in EA groups was higher than that of the same phase model group in 3h～12d,and were superior to that in model groups at 3d and 7d(P<0.05,P<0.01).(2)The mRNA and protein expression of Notch4:the mRNA expression at 3h～12d and protein expression at 9h～12d of rats in model was up-regulated significantly compared to that in control groups(P<0.05,P<0.01).The mRNA levels of rats in EA groups was higher than that of the same phase model group in 1h,12h～12d,and were superior to that in model groups at 12 h,3d and 7d(P<0.05,P<0.01).While the protein levels of rats in EA groups was higher than that of the same phase model group in 1h,9h～12d,and were superior to that in model groups at 12 h,3d and 7d(P<0.05,P<0.01).(3)The mRNA and protein expression of Jagged1:the mRNA expression at 1h～12d and protein expression at 3h～12d of rats in model was up-regulated significantly compared to that in control groups(P<0.05,P<0.01).The mRNA levels of rats in EA groups was higher than that of the same phase model group in 6h,12h～12d,and were superior to that in model groups at 12 h and 24h～12d(P<0.05,P<0.01).While the protein levels of rats in EA groups was higher than that of the same phase model group in 3h～12d,and were superior to that in model groups at 12 h and 7d(P<0.05,P<0.01).(4)The mRNA and protein expression of Dll4:the mRNA expression at 3h～12d and protein expression at 6h～12d of rats in model was up-regulated significantly compared to that in control groups(P<0.05,P<0.01).The mRNA levels of rats in EA groups was higher than that of the same phase model group in 3h,6h,12h～7d,and were superior to that in model groups at 12 h and 24h(P<0.05,P<0.01).While the protein levels of rats in EA groups was higher than that of the same phase model group in 3h～24h and 7d～12d,and were superior to that in model groups at 12 h and 24h(P<0.05,P<0.01).(5)The mRNA and protein expression of hes1:the mRNA expression at 3h～12h,24h～12d and protein expression at 6h～12h,24h～12d of rats in model groups was up-regulated significantly compared to that in control groups(P<0.05,P<0.01).The mRNA levels of rats in EA groups was higher than that of the same phase model group in 3h,6h,18h～7d,and were superior to that in model groups at 7d(P<0.05).While the protein levels of rats in EA groups was higher than that in model group,and were superior to that in model groups at 3d and 7d(P<0.01).Conclusion:1 EA stimulation on DU26 could substantially improve neurological impairment of rats after MCAO,especially at 3d,7d and 12 d after MCAO.2 EA stimulation on DU26 could up-regulate the mRNA and protein expression of VEGF and its receptor VEGFR-1,VEGFR-2 in rat brain tissue after MCAO,participate in the early activation of VEGF signal pathway and angiogenesis after cerebral infarction.3 EA stimulation on DU26 could up-regulate the mRNA and protein expression of vital factors related to Dll4/Notch signaling pathway(Notch1,Notch4,Jagged1,Dll4,hes1),so as to participate in angiogenesis after cerebral infarction in coordination with VEGF signal pathway. |
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